Prevalence of Schistosoma mansoni, soil-transmitted helminths and intestinal protozoa in orphans and street children in Mwanza city, Northern Tanzania.

BACKGROUND: Parasitic infections are highly prevalent in low-income environments worldwide. While orphans and street children represent a particularly vulnerable population group, they are often exempt from preventive interventions such as Mass Drug Administration. In part, this could be due to a lack of data showing the burden of disease in this group. This study aims to address this gap. METHODS: For this cross-sectional study, 144 orphans and 112 street children were screened for Schistosoma mansoni (S. mansoni), Schistosoma haematobium (S. haematobium), soil-transmitted helminths and intestinal protozoa using POC-CCA testing, urine filtration, and Kato-Katz technique. Nutritional status, water- and washing patterns were determined using a standardised questionnaire. Ultrasonography was performed to screen for organ abnormalities. RESULTS: The prevalence of S. mansoni determined by POC-CCA-test was 65.9% for orphans and 94.5% for street children. 19.2% of the orphans tested positive for S. mansoni in Kato Katz. Of the street children, 77.1% showed positive test results in Kato-Katz. Only 1.3% of the orphans stated in the questionnaire that they use the lake to wash, whereas 91.1% of the street children named the lake as at least one of their options for washing. Microscopy showed positive results for Giardia intestinalis (G. intestinalis) in 8.2% and for Entamoeba histolytica/dispar (E. histolytica/dispar) in 23% of orphans and 8.1% for G. intestinalis, and 23.8% for E. histolytica/dispar in street children. In the ultrasonography, we did not observe patterns that indicate severe periportal fibrosis. CONCLUSION: The results indicate a significantly higher rate of infections with S. mansoni in street children compared with orphans. This might be explained by the lack of access to adequate sanitation for street children as well as regular contact with the water of Lake Victoria. However, we did not find similar results concerning infection rates with protozoa. The study results show overall inadequate living conditions in this study population, which could be addressed by public health interventions.

High Egg Reduction Rate but poor clearance of Circulating Cathodic Antigen three weeks after Praziquantel treatment among school children on Ijinga Island, north-western Tanzania.

Traditionally, efficacy of Praziquantel (PZQ) is monitored using Parasitological Cure Rates and Egg Reduction Rates applying Kato Katz (KK) technique. This parasitological technique has a number of limitations. Recently, the Point-of-Care Circulating Cathodic Antigen (POC-CCA) rapid test which is a highly sensitive technique, has emerged as a promising candidate to be used for evaluating the efficacy of PZQ. A prospective longitudinal study was conducted among 399 school children aged 7-17 years on Ijinga Island, north-western Tanzania. At baseline and three weeks after treatment, stool and urine samples were collected from participating school children and screened for S. mansoni infection using the KK technique as well as POC-CCA test. All S. mansoni infected children at baseline were treated with 40mg/kg of PZQ and followed up after three weeks. At baseline, the overall prevalence of S. mansoni infection was 56.6% (95%CI: 51.7-61.4) and 99.7% (95%CI: 98.2-99.9) (considering trace as positive) using KK technique and POC-CCA test, respectively. Three weeks after treatment, the prevalence of S. mansoni was 0.92% using the KK technique and 97.7% when applying the POC-CCA test. The parasitological cure rates based on KK technique and POC-CCA were 99.1% (95%CI: 97.5-99.8) and 2.3% (95%CI: 1.2-4.5). Egg Reduction Rate was 99.1%. Based on WHO guidelines using the KK technique, at three weeks point, the efficacy of PZQ is satisfactory. However, the assessment of the efficacy of PZQ using POC-CCA tests needs further evaluation.

Detection of Schistosoma mansoni DNA using polymerase chain reaction from serum and dried blood spot card samples of an adult population in North-western Tanzania.

BACKGROUND: Real-time polymerase chain reaction (PCR) is a sensitive and specific method for diagnosing schistosomiasis. However, this method should be performed in a laboratory, usually located distant from the sample collection site. Therefore, it is important to have fast sampling preservation methods, which allow simple transport prior to DNA extraction and amplification. The aim of this study was to verify if blood samples applied to filter paper are suitable for analysis of Schistosoma mansoni DNA by real-time PCR. METHODS: A cross-sectional study was conducted among 100 study participants aged 17 to 70 years in a fishing village on the southern shore of Lake Victoria, Tanzania. Serum samples and ethylenediaminetetraacetic acid (EDTA)-anticoagulated whole blood for preparation of dried blood spots (DBS) were collected to test for Schistosoma mansoni infection by real-time PCR. A combined diagnostic reference of positive results of serum-based real-time PCR and the Kato-Katz (KK) method was used for analysis. Sensitivity and negative predictive value (NPV) were calculated. The Wilcoxon signed-rank test was chosen to compare the mean cycle threshold (Ct) values from serum and DBS. RESULTS: According to the reference, 92.5% S. mansoni positive samples were determined. The serum-based real-time PCR performed excellently with 95.4% sensitivity, whereas the DBS-based real-time PCR showed a low sensitivity (45.4%). The Ct-values were significantly higher in DBS (median: 37.3) than in serum samples (median: 27.5, P < 0.001), reflecting a lower parasite-specific DNA load on the filter cards. With increasing egg counts, an increase in sensitivity was observed for all methods. The POC-CCA test and the serum-based real-time PCR showed a sensitivity of 100% for medium and severe infections. The DBS real-time PCR showed a sensitivity of only 85.7% even for severe infections. CONCLUSIONS: DBS-based real-time PCR did not provide good results in our study and therefore should not be recommended or must be tested concerning temperature of storage, storage duration, use of different filter papers and extraction methods before it is used in future studies. In contrast, our results showed that the POC-CCA test is a sensitive and precise test for detecting S. mansoni infections .

Evaluation of serum-based real-time PCR to detect Schistosoma mansoni infection before and after treatment.

BACKGROUND: To detect acute schistosomiasis, low-intensity infections, or to verify the success of treatment with praziquantel, highly sensitive test methods are required. The aim of this study was therefore to demonstrate the performance of Schistosoma mansoni specific DNA detection in serum and urine using real-time polymerase chain reaction (PCR) in an endemic area before and after treatment. METHODS: The study pursued a 1-week and 20-weeks longitudinal design with a treatment intervention among 36 study participants aged 18 to 70 years in the community of Kayenze, a fishing village in Ilemela district on the southern shore of Lake Victoria in north-western Tanzania between February and June 2018. Blood, urine and stool samples were collected from each participant to diagnose Schistosoma mansoni infection before and two times after treatment with praziquantel using serum- and urine based real-time PCR, point-of-care circulating cathodic antigen (POC-CCA) rapid diagnostic test and the microscopic Kato-Katz (KK) method. Kappa coefficient (κ) was used to estimate the agreement between these diagnostic tests compared to a combined "gold standard" of positive results by serum-based real-time PCR and/or positive egg counts determined by KK. Kendall's Tau rank correlation was used to examine the relationship between cycle threshold (Ct)-values and egg counts and the Wilcoxon signed-rank test was used to compare the median Ct-values of the different examination time points. RESULTS: By using the combined "gold standard" of the parasitological Kato-Katz test and/or serum-based real-time PCR, a S. mansoni prevalence of 77.1% could be determined at baseline. In terms of sensitivity, serum-based real-time PCR (96.3%) and POC-CCA assay (77.8%) showed the highest results. The detection of DNA from urine samples showed the lowest sensitivity (33.3%). Treatment with praziquantel resulted in a significantly reduced prevalence of S. mansoni. No infection could be detected by Kato-Katz, with the POC-CCA test only 33.3%. The analysis of the median Ct values over time (which were determined by the serum-based real-time PCR) showed that the Ct decreases significantly shortly after treatment (from 30.3 to 28) and increases above baseline level (34.9) three months later. CONCLUSIONS: The data presented here show that the serum-based real-time PCR exhibits excellent diagnostic accuracy, in contrast to the use of urine as sample material for S. mansoni DNA detection. However, as circulating DNA does not necessarily reflect the persistence of living worms in schistosomiasis, this method is less well suited to verify the success of treatment with praziquantel.

Malacological survey to identify transmission sites for intestinal schistosomiasis on Ijinga Island, Mwanza, north-western Tanzania.

The role of malacological surveys to identify potential transmission sites for schistosomiasis control in this era of mass drug administration have received little attention. In that context, the present study was conducted to determine the abundance, identity and disease transmission potential of intermediate host snails for intestinal schistosomiasis on Ijinga Island, north-western Tanzania. A cross-sectional malacological study was conducted between February and March 2016 on Ijinga Island, Lake Victoria, north-western Tanzania. Snails were collected at points where humans are in frequent contact with water using a standardized scooping technique and have been identified using shell morphological features. The Schistosoma infection status of the collected snails was determined by using real-time Polymerase Chain Reaction (real-time PCR). A total number of 4,888 snails were putatively identified as Biomphalaria species. A random sample of 788 snails underwent molecular analyses for Schistosoma infection. Overall, 279 (35.4%) of Biomphalaria species were identified to be infected with parasites of the lateral spined S. mansoni group. The findings confirm that Biomphalaria species collected in areas with high human water contacts are infected with Schistosoma and that there is a likeliness of local risk for schistosomiasis transmission at most water contact points around Ijinga Island.

Intestinal schistosomiasis of Ijinga Island, north-western Tanzania: prevalence, intensity of infection, hepatosplenic morbidities and their associated factors.

BACKGROUND: Intestinal schistosomiasis is highly endemic in Tanzania and mass drug administration (MDA) using praziquantel is the mainstay of the control program. However, the MDA program covers only school aged children and does not include neither adult individuals nor other public health measures. The Ijinga schistosomiasis project examines the impact of an intensified treatment protocol with praziquantel MDA in combination with additional public health interventions. It aims to investigate the feasibility of eliminating intestinal schistosomiasis in a highly endemic African setting using an integrated community-based approach. In preparation of this project, we report about baseline data on S.mansoni prevalence, intensity of infection, related hepatosplenic morbidities and their associated factors. METHODS: A cross sectional study was conducted among 930 individuals aged 1-95 years living at Ijinga Island, north-western Tanzania in September 2016. Single stool and urine samples were collected from each study participant and processed using Kato Katz (KK) technique and point-of-care Circulating Cathodic (POC-CCA) antigen test for detection of S.mansoni eggs and antigen respectively. Ultrasonographical examination for S.mansoni hepatosplenic morbidities was done to all participants. For statistical analyses Fisher's exact test, chi-square test, student-t-test, ANOVA and linear regression were used where applicable. RESULTS: Overall based on KK technique and POC-CCA test, 68.9% (95%CI: 65.8-71.8) and 94.5% (95%CI: 92.8-95.8) were infected with S.mansoni. The overall geometrical mean eggs per gram (GMepg) of faeces was 85.7epg (95%CI: 77.5-94.8). A total of 27.1, 31.2 and 51.9% of the study participants had periportal fibrosis (PPF-grade C-F), splenomegaly and hepatomegaly. Risk factors for PPF were being male (aRR = 1.08, 95%CI: 1.02-1.16, P < 0.01), belong to the age group 16-25 years (aRR = 1.23, 95%CI: 105-1.44, P < 0.01), 26-35 years (aRR = 1.42, 95%CI: 1.21-1.67, P < 0.001), 36-45 years (aRR = 1.56, 95%CI:1.31-1.84, P < 0.001) and ≥ 46 years (aRR = 1.64, 95%CI:1.41-1.92, P < 0.001). The length of the left liver lobe was associated with being female (P < 0.03), belong to the age group 1-5 years (P < 0.013), 6-15 years (P < 0.04) and S.mansoni intensity of infection (P < 0.034). Male sex (aRR = 1.15, 95%CI:1.06-1.24, P < 0.001) and belonging to the age groups 16-25 years (aRR = 1.27, 95%CI:1.05-1.54, P < 0.02) or 26-35 years (aRR = 1.32, 95%CI:108-1.61, P < 0.01) were associated with splenomegaly. CONCLUSION: Schistosoma mansoni infection and its related morbidities (hepatomegaly, splenomegaly, periportal fibrosis) are common in the study area. Age, sex and intensity of infection were associated with periportal fibrosis. The prevalence of S.mansoni was above 50% in each age group and based on the observed prevalence, we recommend MDA to the entire community.

Comparison of sensitivity and specificity of three diagnostic tests to detect Schistosoma mansoni infections in school children in Mwanza region, Tanzania.

BACKGROUND: Schistosomiasis remains one of the most prevalent parasitic infections in the world and has significant economic and public health consequences, particularly in poor communities. Reliable and accurate diagnosis plays a key role in surveillance, prevention and control of schistosomiasis. Currently, the microscopic Kato Katz (KK) stool thick smear technique is the most commonly used method to diagnose Schistosoma mansoni infections in epidemiological surveys. It is well-known that the sensitivity of this parasitological method decreases when infection intensities are moderate to low, however. The urine-based Point-of Care Circulating Cathodic Antigen (POC-CCA) test has been extensively evaluated as a further diagnostic tool. Several studies have shown that the POC-CCA test is more sensitive but less specific than the KK method. However, to clarify the meaning of inconsistent results between KK and POC-CCA tests in clinical routine, this study compares the accuracy of microscopy and POC-CCA versus real-time polymerase chain reaction (real-time PCR) results of urine and faecal samples from African school children participants. METHODOLOGY: This was a school-based cross-sectional study conducted in 2015 among 305 school children aged 7-16 years from two primary schools located in Ilemela and Magu Districts, north-western Tanzania. Single stool and urine samples were collected from each participant and examined for the presence of Schistosoma mansoni eggs, parasite antigen, and parasite DNA using KK thick smears, POC-CCA tests, and real-time PCR, respectively. PRINCIPAL FINDINGS: The prevalence of S. mansoni infection, calculated by KK was 85.2%, by real-time PCR 92.9% and by POC-CCA 94.9%. In comparison to KK, the POC-CCA and real-time PCR tests had sensitivities of 89.7% and 99.5% and specificities of 22.73% and 29.55%, respectively. However, due to the known limitations of the KK assay, we also used latent class analysis (LCA) that included POC-CCA, KK, and schistosome-specific real-time PCR results to determine their sensitivities and specificities. The POC-CCA test had the highest sensitivity (99.5%) and a specificity of 63.4% by LCA and the real-time PCR test had a sensitivity of 98.7% and the highest specificity (81.2%). CONCLUSION: In moderate and high prevalence areas, the POC-CCA cassette test is more sensitive than the KK method and can be used for screening and geographical mapping of S. mansoni infections. Real-time PCR is highly sensitive and also shows the highest specificity among the 3 investigated diagnostic procedures. It can offer added value in diagnosing schistosomiasis.

Fluorine walk: The impact of fluorine in quinolone amides on their activity against African sleeping sickness.

Human African Trypanosomiasis, also known as African sleeping sickness, is caused by the parasitic protozoa of the genus Trypanosoma. If there is no pharmacological intervention, the parasites can cross the blood-brain barrier (BBB), inevitably leading to death of the patients. Previous investigation identified the quinolone amide GHQ168 as a promising lead compound having a nanomolar activity against T. b. brucei. Here, the role of a fluorine substitution at different positions was investigated in regard to toxicity, pharmacokinetics, and antitrypanosomal activity. This 'fluorine walk' led to new compounds with improved metabolic stability and consistent activity against T. b. brucei. The ability of the new quinolone amides to cross the BBB was confirmed using an 18F-labelled quinolone amide derivative by means of ex vivo autoradiography of a murine brain.

Bistacrines as potential antitrypanosomal agents.

Human African Trypanosomiasis (HAT) is caused by two subspecies of the genus Trypanosoma, namely Trypanosoma brucei rhodesiense and Trypanosoma brucei gambiense. The disease is fatal if left untreated and therapy is limited due to only five non-adequate drugs currently available. In preliminary studies, dimeric tacrine derivatives were found to inhibit parasite growth with IC50-values in the nanomolar concentration range. This prompted the synthesis of a small, but smart library of monomeric and dimeric tacrine-type compounds and their evaluation of antiprotozoal activity. Rhodesain, a lysosomal cathepsin-L like cysteine protease of T. brucei rhodesiense is essential for parasite survival and likely target of the tacrine derivatives. In addition, the inhibition of trypanothione reductase by bistacrines was found. This flavoprotein oxidoreductase is the main defense against oxidative stress in the thiol redox system unique for protozoa.

Anti-trypanosomal activities and structural chemical properties of selected compound classes.

Potent compounds do not necessarily make the best drugs in the market. Consequently, with the aim to describe tools that may be fundamental for refining the screening of candidates for animal and preclinical studies and further development, molecules of different structural classes synthesized within the frame of a broad screening platform were evaluated for their trypanocidal activities, cytotoxicities against murine macrophages J774.1 and selectivity indices, as well as for their ligand efficiencies and structural chemical properties. To advance into their modes of action, we also describe the morphological and ultrastructural changes exerted by selected members of each compound class on the parasite Trypanosoma brucei. Our data suggest that the potential organelles targeted are either the flagellar pocket (compound 77, N-Arylpyridinium salt; 15, amino acid derivative with piperazine moieties), the endoplasmic reticulum membrane systems (37, bisquaternary bisnaphthalimide; 77, N-Arylpyridinium salt; 68, piperidine derivative), or mitochondria and kinetoplasts (88, N-Arylpyridinium salt; 68, piperidine derivative). Amino acid derivatives with fumaric acid and piperazine moieties (4, 15) weakly inhibiting cysteine proteases seem to preferentially target acidic compartments. Our results suggest that ligand efficiency indices may be helpful to learn about the relationship between potency and chemical characteristics of the compounds. Interestingly, the correlations found between the physico-chemical parameters of the selected compounds and those of commercial molecules that target specific organelles indicate that our rationale might be helpful to drive compound design toward high activities and acceptable pharmacokinetic properties for all compound families.